Flavoproteins and flavoenzymes are a ubiquitous class of biological macromolecules involved in an exceedingly wide variety of catalytic and storage functions. These various functions represent many stages in the evolution of flavoproteins. A similar diversity is shown for the NAD-dependent proteins, which have been thoroughly studied by x-ray crystallographic methods. A conclusion drawn from the x-ray studies of the NAD-proteins (See Rossman, et al, 1975) is that all of the NAD proteins have similar spatial configurations around the NAD binding site. Ross and his co-workers (See Subramanian and Ross, 1978) used an elegant calorimetric approach to test the conclusion of Rossman and found a remarkable correlation between the NAD proteins and the thermodynamic parameters for NAD binding. It is currently suggested that the flavoproteins have a substructure which is in many ways similar to the NAD system. The experiments outlined in this proposal are designed to test the hypothesis. In addition, the ionization state of the phosphate of FMN when bound to the proteins will be determined as well as a set of precise thermodynamic parameters for the binding process. The studies will significantly inprove our understanding of the flavin binding process and the flavin binding site as well as provide thermodynamic data to which any proposed flavoenzyme catalytic mechanism may be compared.